U.S. patent number 10,858,307 [Application Number 15/999,358] was granted by the patent office on 2020-12-08 for compositions and methods for treating liver cancer.
This patent grant is currently assigned to Golden Biotechnology Corporation. The grantee listed for this patent is Golden Biotechnology Corporation. Invention is credited to Chih-Ming Chen, Sheng-Yung Liu.
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United States Patent |
10,858,307 |
Liu , et al. |
December 8, 2020 |
Compositions and methods for treating liver cancer
Abstract
Provided herein are compositions and methods of treating liver
cancer by novel anticancer agents.
Inventors: |
Liu; Sheng-Yung (New Taipei,
TW), Chen; Chih-Ming (New Taipei, TW) |
Applicant: |
Name |
City |
State |
Country |
Type |
Golden Biotechnology Corporation |
Jersey City |
NJ |
US |
|
|
Assignee: |
Golden Biotechnology
Corporation (Jersey City, NJ)
|
Family
ID: |
59625477 |
Appl.
No.: |
15/999,358 |
Filed: |
February 16, 2017 |
PCT
Filed: |
February 16, 2017 |
PCT No.: |
PCT/US2017/018236 |
371(c)(1),(2),(4) Date: |
August 17, 2018 |
PCT
Pub. No.: |
WO2017/143106 |
PCT
Pub. Date: |
August 24, 2017 |
Prior Publication Data
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Document
Identifier |
Publication Date |
|
US 20190112258 A1 |
Apr 18, 2019 |
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Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
Issue Date |
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62296097 |
Feb 17, 2016 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C07D
333/20 (20130101); A61P 35/00 (20180101); C07C
49/557 (20130101); C07D 209/14 (20130101); C07D
209/16 (20130101); C07C 225/20 (20130101); C07C
49/543 (20130101); A61K 31/12 (20130101); C07D
213/36 (20130101); C07C 2601/16 (20170501) |
Current International
Class: |
C07C
225/00 (20060101); C07C 225/20 (20060101); C07D
209/14 (20060101); A61P 35/00 (20060101); A61K
31/12 (20060101); C07C 49/543 (20060101); C07D
333/20 (20060101); C07D 213/36 (20060101); C07C
49/557 (20060101); C07D 209/16 (20060101) |
References Cited
[Referenced By]
U.S. Patent Documents
Foreign Patent Documents
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2233463 |
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Sep 2010 |
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EP |
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2233463 |
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Sep 2010 |
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EP |
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Other References
Han, H. "Targeted Prodrug Design to Optimize Drug Delivery." AAPS
Pharmsci. (2000), vol. 2 (1) article 6, pp. 1-11. (Year: 2000).
cited by examiner .
Ettmayer, P., et al. "Lessons Learned from Marketed and
Investigational Prodrugs." J. Med. Chem. (2004) 47(10), pp.
2393-2404. (Year: 2004). cited by examiner .
Testa, B. "Prodrug research: futile or fertile?" Biochem. Pharm.
(2004) 68, pp. 2097-2106. (Year: 2004). cited by examiner.
|
Primary Examiner: Kenyon; John S
Attorney, Agent or Firm: Liang; Chang-Hsing
Claims
What is claimed is:
1. A compound of formula I: ##STR00033## or a pharmaceutically
acceptable salt, metabolite, or solvate thereof, wherein each of R,
and R.sup.1 independently is a hydrogen, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl
optionally substituted with one or more halogen, NR.sub.5R.sub.6,
OR.sub.5, aryl or heteroaryl; R.sup.2 is a hydrogen,
C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, or
C.sub.1-C.sub.8alkyl optionally substituted with one or more
halogen, aryl or heteroaryl, R.sup.3 is a hydrogen, optionally
substituted methyl or (CH.sub.2).sub.m--CH.sub.3, m=0-6, R.sup.4 is
H, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6,
C.sub.1-C.sub.8alkyl, C.sub.2-C.sub.8alkenyl,
C.sub.2-C.sub.8alkynyl, wherein the C.sub.1-C.sub.8alkyl,
C.sub.2-C.sub.8alkenyl, C.sub.2-C.sub.8alkynyl, are optionally
substituted with one or more substituents selected from
NR.sub.5R.sub.6, OR.sub.5, OC(.dbd.O)R.sub.7, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, C.sub.1-C.sub.8 alkyl,
C.sub.2-C.sub.8 alkenyl, C.sub.2-C.sub.8 alkynyl, C.sub.3-C.sub.8
cycloalkyl, and C.sub.1-C.sub.8 haloalkyl; each of R.sub.5 and
R.sub.6 is independently H or C.sub.1-C.sub.8alkyl; R.sub.7 is a
C.sub.1-C.sub.8alkyl, OR.sub.5 or NR.sub.5R.sub.6.
2. The compound of claim 1, wherein R.sup.3 is a methyl and R.sup.4
is H.
3. The compound of claim 1, wherein R.sup.3 is hydrogen, methyl,
ethyl, propyl, butyl, pentyl, hexyl.
4. The compound of claim 1, wherein R.sup.4 is H methyl, ethyl,
propyl, butyl, pentyl, hexyl, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
or C(.dbd.O)NR.sub.5R.sub.6.
5. The compound of claim 1, wherein each of R, and R.sup.1
independently is a hydrogen, methyl, ethyl, propyl, butyl, pentyl,
hexyl optionally substituted with one or more halogen,
NR.sub.5R.sub.6, OR.sub.5, aryl or heteroaryl.
6. The compound of claim 5, wherein R.sup.1 is a hydrogen, methyl,
ethyl, propyl, butyl, pentyl, hexyl optionally substituted with one
or more halogen, NR.sub.5R.sub.6, or heteroaryl.
7. The compound of claim 1, wherein R is hydrogen, or
C.sub.1-C.sub.8alkyl.
8. The compound of claim 1, R is hydrogen, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, or C(.dbd.O)NR.sub.5R.sub.6.
9. The compound of claim 1, wherein R.sup.1 is hydrogen, or
C.sub.1-C.sub.8alkyl optionally substituted with one or more
halogen, NR.sub.5R.sub.6, OR.sub.5, aryl or heteroaryl, and R is H,
or C(.dbd.O)R.sub.5.
10. The compound of claim 9, wherein R.sup.2 is methyl, ethyl,
propyl, butyl, pentyl, hexyl.
11. The compound of claim 9, wherein R.sup.2 is C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6.
12. The compound of claim 1, wherein said compound is selected from
the group consisting of ##STR00034## ##STR00035##
13. The compound of claim 1, wherein said compound is selected from
the group consisting of ##STR00036##
14. A method for treating liver cancer in a subject comprising
administering to the subject an effective amount of a compound of
claim 1.
15. The method of claim 14, wherein said compound inhibits liver
cancer cells.
16. The method of claim 14, wherein said compound decreases cancer
tumor mass weight.
17. The method of claim 14, wherein said compound reduces cancer
tumor size.
18. The method of claim 14, wherein said compound is selected from
the group consisting of ##STR00037##
Description
BACKGROUND OF THE INVENTION
Liver cancer, also known as hepatic cancer, is a cancer that
originates in the liver. Primary liver cancer is globally the sixth
most frequent cancer, and the second leading cause of cancer death.
The most frequent liver cancer, accounting for approximately 75% of
all primary liver cancers, is hepatocellular carcinoma (HCC) (also
named hepatoma, which is a misnomer because adenomas are usually
benign). HCC is a cancer formed by liver cells, known as
hepatocytes that become malignant. Hepatocellular carcinoma
accounts for more than half million deaths annually worldwide.
SUMMARY OF THE INVENTION
In one aspect, there are provided a compound of formula I:
##STR00001## or a pharmaceutically acceptable salt, metabolite,
solvate or prodrug thereof, wherein each of each of R, and R.sup.1
independently is a hydrogen, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl optionally
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
aryl or heteroaryl; R.sup.2 is a hydrogen, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl
optionally substituted with one or more halogen, aryl or
heteroaryl, R.sup.3 is a hydrogen, optionally substituted methyl or
(CH.sub.2).sub.m--CH.sub.3, R.sup.4 is H, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, C.sub.1-C.sub.8alkyl,
C.sub.2-C.sub.8alkenyl, C.sub.2-C.sub.8alkynyl, wherein the
C.sub.1-C.sub.5alkyl, C.sub.2-C.sub.8alkenyl,
C.sub.2-C.sub.8alkynyl, are optionally substituted with one or more
substituents selected from NR.sub.5R.sub.6, OR.sub.5,
OC(.dbd.O)R.sub.7, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, C.sub.1-C.sub.8 alkyl, C.sub.2-C.sub.8
alkenyl, C.sub.2-C.sub.8 alkynyl, C.sub.3-C.sub.8 cycloalkyl, and
C.sub.1-C.sub.8 haloalkyl; each of R.sub.5 and R.sub.6 is
independently H or C.sub.1-C.sub.5alkyl; R.sub.7 is a
C.sub.1-C.sub.5alkyl, OR.sub.5 or NR.sub.5R.sub.6.
In one aspect, there are provided a compound of formula Ia:
##STR00002## or a pharmaceutically acceptable salt, metabolite,
solvate or prodrug thereof, wherein each of each of R, and R.sup.1
independently is a hydrogen, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl optionally
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
aryl or heteroaryl; R.sup.2 is a hydrogen, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl
optionally substituted with one or more halogen, aryl or
heteroaryl, and each of R.sub.5 and R.sub.6 is independently H or
C.sub.1-C.sub.8alkyl.
In another aspect, there are provided methods for treating or
reducing the risk of liver cancer in a subject comprising
administering to said subject a therapeutically effective amount of
an anti-cancer agent having the structure:
##STR00003## or a pharmaceutically acceptable salt, metabolite,
solvate or prodrug thereof, wherein each of each of R, and R.sup.1
independently is a hydrogen, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl optionally
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
aryl or heteroaryl; R.sup.2 is a hydrogen, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl
optionally substituted with one or more halogen, NR.sub.5R.sub.6,
OR.sub.5, C.sub.6-C.sub.10aryl or C.sub.4-C.sub.10heteroaryl,
R.sup.3 is a hydrogen, optionally substituted methyl or
(CH.sub.2).sub.m--CH.sub.3, R.sup.4 is H, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, C.sub.1-C.sub.8alkyl,
C.sub.2-C.sub.8alkenyl, C.sub.2-C.sub.8alkynyl, wherein the
C.sub.1-C.sub.8alkyl, C.sub.2-C.sub.8alkenyl,
C.sub.2-C.sub.8alkynyl, are optionally substituted with one or more
substituents selected from NR.sub.5R.sub.6, OR.sub.5,
OC(.dbd.O)R.sub.7, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, C.sub.1-C.sub.8 alkyl, C.sub.2-C.sub.8
alkenyl, C.sub.2-C.sub.8 alkynyl, C.sub.3-C.sub.8 cycloalkyl, and
C.sub.1-C.sub.8 haloalkyl; each of R.sub.5 and R.sub.6 is
independently H or C.sub.1-C.sub.8alkyl; and R.sub.7 is a
C.sub.1-C.sub.8alkyl, OR.sub.5 or NR.sub.5R.sub.6.
INCORPORATION BY REFERENCE
All publications, patents, and patent applications mentioned in
this specification are herein incorporated by reference to the same
extent as if each individual publication, patent, or patent
application was specifically and individually indicated to be
incorporated by reference.
BRIEF DESCRIPTION OF THE DRAWINGS
The novel features of the invention are set forth with
particularity in the appended claims. A better understanding of the
features and advantages of the present invention will be obtained
by reference to the following detailed description that sets forth
illustrative embodiments, in which the principles of the invention
are utilized, and the accompanying drawings of which:
FIG. 1 illustrates the tumor size test results of THP-1 xenograft
mice treated with and without the exemplary invention compound.
FIG. 2 illustrates the tumor mass test results by weight of THP-1
xenograft mice treated with and without with the exemplary
invention compound.
FIG. 3 illustrates the body weight test results of THP-1 xenograft
mice treated with and without the exemplary invention compound.
DETAILED DESCRIPTION OF THE INVENTION
The present invention provides novel anticancer agents and methods
of treating liver cancer therefrom. For example, the following
exemplary anticancer compounds 1-11 were prepared and test for
anticancer activities over liver cancer cells. The exemplary
anticancer agents are analogs of
4-hydroxy-2,3-dimethoxy-6-methyl-5-(3,7,11-trimethyldodeca-2,6,10-trienyl-
)cyclohex-2-enone (Compound S). However, it was found unexpectedly
that those 3-amino derivatives possess superb inhibition, compared
to their 3-hydroxyl analog,
4-hydroxy-2,3-dimethoxy-6-methyl-5-(3,7,11-trimethyldodeca-2,6,10-trienyl-
)cyclohex-2-enone, against liver cancer cells with some of them 10
to 20 folds better. Subsequent in vivo study confirms these
compounds are good candidate for liver cancer treatment.
##STR00004## ##STR00005##
In some embodiments, there are provided herein a compound of
formula I
##STR00006## or a pharmaceutically acceptable salt, metabolite,
solvate or prodrug thereof, wherein each of R, and R.sup.1
independently is a hydrogen, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.5alkyl optionally
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
aryl or heteroaryl; R.sup.2 is a hydrogen, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl
optionally substituted with one or more halogen, aryl or
heteroaryl, R.sup.3 is a hydrogen, optionally substituted methyl or
(CH.sub.2).sub.m--CH.sub.3, R.sup.4 is H, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, C.sub.1-C.sub.8alkyl,
C.sub.2-C.sub.8alkenyl, C.sub.2-C.sub.8alkynyl, wherein the
C.sub.1-C.sub.8alkyl, C.sub.2-C.sub.8alkenyl,
C.sub.2-C.sub.8alkynyl, are optionally substituted with one or more
substituents selected from NR.sub.5R.sub.6, OR.sub.5,
OC(.dbd.O)R.sub.7, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, C.sub.1-C.sub.8 alkyl, C.sub.2-C.sub.8
alkenyl, C.sub.2-C.sub.8 alkynyl, C.sub.3-C.sub.8 cycloalkyl, and
C.sub.1-C.sub.8 haloalkyl; each of R.sub.5 and R.sub.6 is
independently H or C.sub.1-C.sub.8alkyl; and R.sub.7 is a
C.sub.1-C.sub.5alkyl, OR.sub.5 or NR.sub.5R.sub.6.
In certain embodiments, R is hydrogen (H), C.sub.1-C.sub.8alkyl, or
C.sub.1-C.sub.8alkyl substituted with one or more halogen,
NR.sub.5R.sub.6, OR.sub.5, C.sub.6-C.sub.10aryl or
C.sub.4-C.sub.10heteroaryl. In certain embodiments, R is a hydrogen
(H), methyl, ethyl, propyl, butyl, pentyl, hexyl, or the like,
optionally substituted with one or more halogen, NR.sub.5R.sub.6,
OR.sub.5, C.sub.6-C.sub.10aryl or C.sub.4-C.sub.10heteroaryl. In
certain embodiments, R is H, or C.sub.1-C.sub.5alkyl. In certain
embodiments, R is hydrogen, methyl, ethyl, propyl, butyl, pentyl,
or hexyl. In certain embodiments, R is a hydrogen,
C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5, or C(.dbd.O)NR.sub.5R.sub.6.
In certain embodiments, R is H, C.sub.1-C.sub.8alkyl,
C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5, or C(.dbd.O)NR.sub.5R.sub.6.
In certain embodiments, R is H, or C(.dbd.O)R.sub.5. In certain
embodiments, R is H, C(.dbd.O)C.sub.3H.sub.8,
C(.dbd.O)C.sub.2H.sub.5, or C(.dbd.O)CH.sub.3. In certain
embodiments, R is H, C.sub.1-C.sub.8alkyl, or C.sub.1-C.sub.8alkyl
substituted with one or more NR.sub.5R.sub.6, or
C.sub.4-C.sub.10heteroaryl.
In certain embodiments, R.sup.1 is hydrogen (H). In certain
embodiments, R.sup.1 is hydrogen, C.sub.1-C.sub.8alkyl, or
C.sub.1-C.sub.8alkyl substituted with one or more halogen,
NR.sub.5R.sub.6, OR.sub.5, C.sub.6-C.sub.10aryl or
C.sub.4-C.sub.10heteroaryl. In certain embodiments,
C.sub.6-C.sub.10aryl is phenyl. In certain embodiments,
C.sub.4-C.sub.10heteroaryl is pyridyl, thiophenyl, or indolyl. In
certain embodiments, R.sup.1 is hydrogen, or C.sub.1-C.sub.8alkyl.
In certain embodiments, R.sup.1 is H, C.sub.1-C.sub.8alkyl, or
C.sub.1-C.sub.8alkyl substituted with one or more NR.sub.5R.sub.6,
or C.sub.4-C.sub.10heteroaryl. In certain embodiments, R.sup.1 is a
hydrogen (H), methyl, ethyl, propyl, butyl, pentyl, hexyl, or the
like, optionally substituted with one or more halogen,
NR.sub.5R.sub.6, OR.sub.5, C.sub.6-C.sub.10aryl or
C.sub.4-C.sub.10heteroaryl. In certain embodiments, R.sup.1 is
hydrogen, methyl, ethyl, propyl, butyl, optionally substituted with
NR.sub.5R.sub.6, OR.sub.5, C.sub.6-C.sub.10aryl or
C.sub.4-C.sub.10heteroaryl. In certain embodiments, R.sup.1 is
hydrogen, methyl, methyl substituted with
C.sub.4-C.sub.10heteroaryl such as thiophenyl, ethyl substituted
with OR.sub.5 such as OH, ethyl substituted with
C.sub.4-C.sub.10heteroaryl such as indolyl, propyl substituted with
NR.sub.5R.sub.6 such as NH.sub.2. In certain embodiments, R.sup.1
is hydrogen, methyl, methyl substituted with
C.sub.4-C.sub.10heteroaryl such as thiophenyl, or propyl
substituted with NR.sub.5R.sub.6 such as NH.sub.2.
In some embodiments, R is hydrogen, or C(.dbd.O)R.sub.5; and
R.sup.1 is hydrogen, C.sub.1-C.sub.8alkyl, or C.sub.1-C.sub.8alkyl
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
C.sub.6-C.sub.10aryl or C.sub.4-C.sub.10heteroaryl. In certain
embodiments, R is C.sub.1-C.sub.8alkyl and R.sup.1 is
C.sub.1-C.sub.8alkyl. In certain embodiments, R is hydrogen, and
R.sup.1 is hydrogen, C.sub.1-C.sub.8alkyl, or C.sub.1-C.sub.8alkyl
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
C.sub.6-C.sub.10aryl or C.sub.4-C.sub.10heteroaryl. In particular,
R is hydrogen, and R.sup.1 is hydrogen, methyl, ethyl, propyl,
butyl, optionally substituted with NR.sub.5R.sub.6, OR.sub.5,
C.sub.6-C.sub.10aryl or C.sub.4-C.sub.10heteroaryl. In certain
embodiments, R is hydrogen, and R.sup.1 is hydrogen, methyl, methyl
substituted with C.sub.4-C.sub.10heteroaryl such as thiophenyl,
ethyl substituted with OR.sub.5 such as OH, ethyl substituted with
C.sub.4-C.sub.10heteroaryl such as indolyl, or propyl substituted
with NR.sub.5R.sub.6 such as NH.sub.2. In certain embodiments, R is
hydrogen or methyl, and R.sup.1 is hydrogen, methyl, methyl
substituted with C.sub.4-C.sub.10heteroaryl such as thiophenyl, or
propyl substituted with NR.sub.5R.sub.6 such as NH.sub.2.
In some embodiments, R.sup.2 is methyl, ethyl, propyl, butyl,
pentyl, hexyl, or the like. In some embodiments, R.sup.2 is
C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6. In
some embodiments, R.sup.2 is C(.dbd.O)C.sub.3H.sub.7,
C(.dbd.O)C.sub.2H.sub.5, C(.dbd.O)CH.sub.3,
C(.dbd.O)OC.sub.3H.sub.7, C(.dbd.O)OC.sub.2H.sub.5,
C(.dbd.O)OCH.sub.3, C(.dbd.O)NHC.sub.3H.sub.7,
C(.dbd.O)NHC.sub.2H.sub.5, or C(.dbd.O)NHCH.sub.3. In certain
embodiments, R.sup.2 is methyl.
In some embodiments, R.sup.3 is hydrogen, methyl, ethyl, propyl,
butyl, pentyl, hexyl, or the like. In certain embodiments, R.sup.3
is methyl.
In some embodiments, R.sup.4 is hydrogen (H), C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, or
C.sub.1-C.sub.8alkyl. In certain embodiments, R.sup.4 is H
hydrogen, methyl, ethyl, propyl, butyl, pentyl, hexyl, or the like.
In some embodiments, R.sup.4 is C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6. In some embodiments,
R.sup.4 is C(.dbd.O)C.sub.3H.sub.7, C(.dbd.O)C.sub.2H.sub.5,
C(.dbd.O)CH.sub.3, C(.dbd.O)OC.sub.3H.sub.7,
C(.dbd.O)OC.sub.2H.sub.5, C(.dbd.O)OCH.sub.3,
C(.dbd.O)NHC.sub.3H.sub.7, C(.dbd.O)NHC.sub.2H.sub.5, or
C(.dbd.O)NHCH.sub.3. In certain embodiments, R.sup.4 is
hydrogen.
In some embodiments, there are provided herein a compound of
formula Ia
##STR00007## or a pharmaceutically acceptable salt, metabolite,
solvate or prodrug thereof, wherein each of each of R, and R.sup.1
independently is a hydrogen, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl optionally
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
aryl or heteroaryl; R.sup.2 is a hydrogen, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl
optionally substituted with one or more halogen, aryl or
heteroaryl; and each of R.sub.5 and R.sub.6 is independently H or
C.sub.1-C.sub.8alkyl.
In some embodiments, each of R, and R.sup.1 independently is a
hydrogen, methyl, ethyl, propyl, butyl, pentyl, hexyl, or the like,
optionally substituted with one or more halogen, NR.sub.5R.sub.6,
OR.sub.5, aryl or heteroaryl. In certain embodiments, R is H,
C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6. In
certain embodiments, R is H, or C(.dbd.O)R.sub.5. In certain
embodiments, R is H, C(.dbd.O)C.sub.3H, C(.dbd.O)C.sub.2H.sub.5, or
C(.dbd.O)CH.sub.3. In certain embodiments, R.sup.1 is hydrogen. In
certain embodiments, R is hydrogen, C.sub.1-C.sub.8alkyl,
C.sub.1-C.sub.8alkyl substituted with one or more halogen,
NR.sub.5R.sub.6, OR.sub.5, aryl or heteroaryl. In certain
embodiments, aryl is phenyl. In certain embodiments, heteroaryl is
pyridyl, thiophenyl, or indolyl.
In some embodiments, R.sup.2 is methyl, ethyl, propyl, butyl,
pentyl, hexyl, or the like. In some embodiments, R.sup.2 is
C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6. In
some embodiments, R.sup.2 is C(.dbd.O)C.sub.3H.sub.7,
C(.dbd.O)C.sub.2H.sub.5, C(.dbd.O)CH.sub.3,
C(.dbd.O)OC.sub.3H.sub.7, C(.dbd.O)OC.sub.2H.sub.5,
C(.dbd.O)OCH.sub.3, C(.dbd.O)NHC.sub.3H.sub.7,
C(.dbd.O)NHC.sub.2H.sub.5, or C(.dbd.O)NHCH.sub.3. In certain
embodiments, R.sup.2 is methyl.
In certain embodiments provide a compound of formula I
##STR00008## or a pharmaceutically acceptable salt, metabolite,
solvate or prodrug thereof, wherein the compound is selected from
the group consisting of
##STR00009## ##STR00010##
In certain embodiments, the compound is selected from the group
consisting of
##STR00011##
The following are some non-limited examples of invention compounds
useful for anticancer treatments:
##STR00012## ##STR00013## ##STR00014##
In some embodiments, the compounds disclosed herein are useful for
treating or reducing the risk of liver cancer in a subject,
especially in view of their unexpected benefit in inhibition of
liver cancer cells in comparison with their respective 3-hydroxyl
analogs.
In accordance with the present invention, there are provided
methods for treating or reducing the risk of liver cancer in a
subject comprising administering to said subject a therapeutically
effective amount of an anti-cancer agent having the structure:
##STR00015## or a pharmaceutically acceptable salt, metabolite,
solvate or prodrug thereof, wherein each of each of R, and R.sup.1
independently is a hydrogen, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, or C.sub.1-C.sub.8alkyl optionally
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
aryl or heteroaryl; each of R.sup.2, and R.sup.3 independently is a
hydrogen, optionally substituted methyl or
(CH.sub.2).sub.m--CH.sub.3, R.sub.4 is H, C(.dbd.O)OR.sub.5,
C(.dbd.O)R.sub.5, C(.dbd.O)NR.sub.5R.sub.6, C.sub.1-C.sub.8alkyl,
C.sub.2-C.sub.8alkenyl, C.sub.2-C.sub.8alkynyl, wherein the
C.sub.1-C.sub.8alkyl, C.sub.2-C.sub.8alkenyl,
C.sub.2-C.sub.8alkynyl, are optionally substituted with one or more
substituents selected from NR.sub.5R.sub.6, OR.sub.5,
OC(.dbd.O)R.sub.7, C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5,
C(.dbd.O)NR.sub.5R.sub.6, C.sub.1-C.sub.8 alkyl, C.sub.2-C.sub.8
alkenyl, C.sub.2-C.sub.8 alkynyl, C.sub.3-C.sub.8 cycloalkyl, and
C.sub.1-C.sub.8 haloalkyl; each of R.sub.5 and R.sub.6 is
independently H or C.sub.1-C.sub.8alkyl; R.sub.7 is a
C.sub.1-C.sub.8alkyl, OR.sub.5 or NR.sub.5R.sub.6.
In certain embodiments, R is hydrogen (H), C.sub.1-C.sub.8alkyl, or
C.sub.1-C.sub.8alkyl substituted with one or more halogen,
NR.sub.5R.sub.6, OR.sub.5, C.sub.6-C.sub.10aryl or
C.sub.4-C.sub.10heteroaryl. In certain embodiments, R is a hydrogen
(H), methyl, ethyl, propyl, butyl, pentyl, hexyl, or the like,
optionally substituted with one or more halogen, NR.sub.5R.sub.6,
OR.sub.5, C.sub.6-C.sub.10aryl or C.sub.4-C.sub.10heteroaryl. In
certain embodiments, R is H, or C.sub.1-C.sub.8alkyl. In certain
embodiments, R is hydrogen, methyl, ethyl, propyl, butyl, pentyl,
or hexyl. In certain embodiments, R is a hydrogen,
C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5, or C(.dbd.O)NR.sub.5R.sub.6.
In certain embodiments, R is H, C.sub.1-C.sub.8alkyl,
C(.dbd.O)OR.sub.5, C(.dbd.O)R.sub.5, or C(.dbd.O)NR.sub.5R.sub.6.
In certain embodiments, R is H, or C(.dbd.O)R.sub.5. In certain
embodiments, R is H, C(.dbd.O)C.sub.3H.sub.8,
C(.dbd.O)C.sub.2H.sub.5, or C(.dbd.O)CH.sub.3. In certain
embodiments, R is H, C.sub.1-C.sub.8alkyl, or C.sub.1-C.sub.8alkyl
substituted with one or more NR.sub.5R.sub.6, or
C.sub.4-C.sub.10heteroaryl.
In certain embodiments, R.sup.1 is hydrogen (H). In certain
embodiments, R.sup.1 is hydrogen, C.sub.1-C.sub.8alkyl, or
C.sub.1-C.sub.8alkyl substituted with one or more halogen,
NR.sub.5R.sub.6, OR.sub.5, C.sub.6-C.sub.10aryl or
C.sub.4-C.sub.10heteroaryl. In certain embodiments,
C.sub.6-C.sub.10aryl is phenyl. In certain embodiments,
C.sub.4-C.sub.10heteroaryl is pyridyl, thiophenyl, or indolyl. In
certain embodiments, R.sup.1 is hydrogen, or C.sub.1-C.sub.8alkyl.
In certain embodiments, R.sup.1 is H, C.sub.1-C.sub.8alkyl, or
C.sub.1-C.sub.8alkyl substituted with one or more NR.sub.5R.sub.6,
or C.sub.4-C.sub.10heteroaryl. In certain embodiments, R.sup.1 is a
hydrogen (H), methyl, ethyl, propyl, butyl, pentyl, hexyl, or the
like, optionally substituted with one or more halogen,
NR.sub.5R.sub.6, OR.sub.5, C.sub.6-C.sub.10aryl or
C.sub.4-C.sub.10heteroaryl. In certain embodiments, R.sup.1 is
hydrogen, methyl, ethyl, propyl, butyl, optionally substituted with
NR.sub.5R.sub.6, OR.sub.5, C.sub.6-C.sub.10aryl or
C.sub.4-C.sub.10heteroaryl. In certain embodiments, R.sup.1 is
hydrogen, methyl, methyl substituted with
C.sub.4-C.sub.10heteroaryl such as thiophenyl, ethyl substituted
with OR.sub.5 such as OH, ethyl substituted with
C.sub.4-C.sub.10heteroaryl such as indolyl, propyl substituted with
NR.sub.5R.sub.6 such as NH.sub.2. In certain embodiments, R.sup.1
is hydrogen, methyl, methyl substituted with
C.sub.4-C.sub.10heteroaryl such as thiophenyl, or propyl
substituted with NR.sub.5R.sub.6 such as NH.sub.2.
In some embodiments, R is hydrogen, or C(.dbd.O)R.sub.5; and
R.sup.1 is hydrogen, C.sub.1-C.sub.8alkyl, or C.sub.1-C.sub.8alkyl
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
C.sub.6-C.sub.10aryl or C.sub.4-C.sub.10heteroaryl. In certain
embodiments, R is C.sub.1-C.sub.8alkyl and R.sup.1 is
C.sub.1-C.sub.8alkyl. In certain embodiments, R is hydrogen, and
R.sup.1 is hydrogen, C.sub.1-C.sub.8alkyl, or C.sub.1-C.sub.8alkyl
substituted with one or more halogen, NR.sub.5R.sub.6, OR.sub.5,
C.sub.6-C.sub.10aryl or C.sub.4-C.sub.10heteroaryl. In particular,
R is hydrogen, and R.sup.1 is hydrogen, methyl, ethyl, propyl,
butyl, optionally substituted with NR.sub.5R.sub.6, OR.sub.5,
C.sub.6-C.sub.10aryl or C.sub.4-C.sub.10heteroaryl. In certain
embodiments, R is hydrogen, and R.sup.1 is hydrogen, methyl, methyl
substituted with C.sub.4-C.sub.10heteroaryl such as thiophenyl,
ethyl substituted with OR.sub.5 such as OH, ethyl substituted with
C.sub.4-C.sub.10heteroaryl such as indolyl, or propyl substituted
with NR.sub.5R.sub.6 such as NH.sub.2. In certain embodiments, R is
hydrogen or methyl, and R.sup.1 is hydrogen, methyl, methyl
substituted with C.sub.4-C.sub.10heteroaryl such as thiophenyl, or
propyl substituted with NR.sub.5R.sub.6 such as NH.sub.2.
It is to be understood that the foregoing general description and
the following detailed description are exemplary and explanatory
only and are not restrictive of any subject matter claimed. In this
application, the use of the singular includes the plural unless
specifically stated otherwise. It must be noted that, as used in
the specification and the appended claims, the singular forms "a,"
"an" and "the" include plural referents unless the context clearly
dictates otherwise. In this application, the use of "or" means
"and/or" unless stated otherwise. Furthermore, use of the term
"including" as well as other forms, such as "include", "includes,"
and "included," is not limiting.
Unless otherwise indicated, conventional methods of mass
spectroscopy, NMR, HPLC, protein chemistry, biochemistry,
recombinant DNA techniques and pharmacology are employed. Unless
specific definitions are provided, the standard nomenclature
employed in connection with, and the standard laboratory procedures
and techniques of, analytical chemistry, synthetic organic
chemistry, and medicinal and pharmaceutical chemistry are employed.
In certain instances, standard techniques are used for chemical
syntheses, chemical analyses, pharmaceutical preparation,
formulation, and delivery, and treatment of patients. In certain
embodiments, standard techniques are used for recombinant DNA,
oligonucleotide synthesis, and tissue culture and transformation
(e.g., electroporation, lipofection). In some embodiments,
reactions and purification techniques are performed e.g., using
kits of manufacturer's specifications or as commonly accomplished
or as described herein.
As used throughout this application and the appended claims, the
following terms have the following meanings:
The term "alkyl" as used herein, means a straight, branched chain,
or cyclic (in this case, it would also be known as "cycloalkyl")
hydrocarbon containing from 1-10 carbon atoms. Illustrative
examples of alkyl include, but are not limited to, methyl, ethyl,
n-propyl, iso-propyl, n-butyl, sec-butyl, tert-butyl, n-pentyl,
isopentyl, neopentyl, n-hexyl, 3-methylhexyl, 2,2-dimethylpentyl,
2,3-dimethylhexyl, n-heptyl, n-octyl, n-nonyl, and n-decyl.
The term "C.sub.1-Calkyl" as used herein, means a straight,
branched chain, or cyclic (in this case, it would also be known as
"cycloalkyl") hydrocarbon containing from 1-8 carbon atoms.
Representative examples of alkyl include, but are not limited to,
methyl, ethyl, n-propyl, iso-propyl, cyclopyl, n-butyl, sec-butyl,
tert-butyl, cyclobutyl, n-pentyl, isopentyl, neopentyl,
cyclopentyl, and n-hexyl.
The term "halo" or "halogen" as used herein, means a --Cl, --Br,
--I or --F.
As used herein, the term "aryl" refers to an aromatic ring wherein
each of the atoms forming the ring is a carbon atom. Aryl rings are
formed by five, six, seven, eight, nine, or more than nine carbon
atoms. Aryl groups are optionally substituted. In one aspect, an
aryl is a phenyl or a naphthalenyl. In one aspect, an aryl is a
phenyl. In one aspect, an aryl is a C.sub.6-C.sub.10aryl. Depending
on the structure, an aryl group can be a monoradical or a diradical
(i.e., an arylene group). In one aspect, an arylene is a
C.sub.6-C.sub.10 arylene. Exemplary arylenes include, but are not
limited to, phenyl-1,2-ene, phenyl-1,3-ene, and phenyl-1,4-ene.
As used herein, the term "heteroaryl" or
"C.sub.4-C.sub.10heteroaryl" employed alone or in combination with
other terms, refers to a monocyclic or polycyclic (e.g., having 2,
3 or 4 fused rings) aromatic hydrocarbon moiety, having one or more
heteroatom ring members selected from nitrogen, sulfur and oxygen,
and having 4 to 10 carbon atoms. In some embodiments, the
heteroaryl group has 1, 2, 3, or 4 heteroatoms. In some
embodiments, the heteroaryl group has 1, 2, or 3 heteroatoms. In
some embodiments, the heteroaryl group has 1 or 2 heteroatoms. In
some embodiments, the heteroaryl group has 1 heteroatom. When the
heteroaryl group contains more than one heteroatom ring member, the
heteroatoms may be the same or different.
Example heteroaryl groups include, but are not limited to,
pyrrolyl, azolyl, oxazolyl, thiazolyl, imidazolyl, furyl, thienyl,
quinolinyl, isoquinolinyl, indolyl, benzothienyl, benzofuranyl,
benzisoxazolyl, imidazo[1,2-b]thiazolyl or the like. In certain
embodiments, the heteroaryl group has 5 to 10 carbon atoms.
The term "optionally substituted" or "substituted" means that the
referenced group may be substituted with one or more additional
group(s) indicated thereof. In some embodiments, if no additional
group(s) indicated thereof, the group(s) may be individually and
independently selected from alkyl, cycloalkyl, aryl, heteroaryl,
heteroalicyclic, hydroxy, alkoxy, aryloxy, alkylthio, arylthio,
alkylsulfoxide, arylsulfoxide, alkylsulfone, arylsulfone, cyano,
halo, nitro, haloalkyl, fluoroalkyl, fluoroalkoxy, and amino,
including mono- and di-substituted amino groups, and the protected
derivatives thereof. By way of example an optional substituents may
be halide, --CN, --NO.sub.2, or L.sub.sR.sub.s, wherein each
L.sub.s is independently selected from a bond, --O--,
--C(.dbd.O)--, --C(.dbd.O)O--, --S--, --S(.dbd.O)--,
--S(.dbd.O).sub.2--, --NH--, --NHC(.dbd.O)--, --C(.dbd.O)NH--,
S(.dbd.O).sub.2NH--, --NHS(.dbd.O).sub.2, --OC(.dbd.O)NH--,
--NHC(.dbd.O)O--, or --(C.sub.1-C.sub.6 alkylene)-; and each
R.sub.s is selected from H, alkyl, fluoroalkyl, heteroalkyl,
cycloalkyl, aryl, heteroaryl, or heterocycloalkyl. The protecting
groups that may form the protective derivatives of the above
substituents may be found in sources such as Greene and Wuts,
above. In some embodiments, optional substituents are selected from
halogen, --CN, --NH.sub.2, --OH, --N(CH.sub.3).sub.2, alkyl,
fluoroalkyl, heteroalkyl, cycloalkyl, heterocycloalkyl, aryl,
heteroaryl, alkoxy, aryloxy, alkylthio, arylthio, alkylsulfoxide,
arylsulfoxide, alkylsulfone, and arylsulfone. In some embodiments,
an optional substituents is halogen, --CN, --NH.sub.2, --OH,
--NH(CH.sub.3), --N(CH.sub.3).sub.2, alkyl, fluoroalkyl,
heteroalkyl, alkoxy, fluoroalkoxy, --S-alkyl, or
--S(.dbd.O).sub.2alkyl. In some embodiments, an optional
substituent is selected from halogen, --CN, --NH.sub.2, --OH,
--NH(CH.sub.3), --N(CH.sub.3).sub.2, --CH.sub.3,
--CH.sub.2CH.sub.3, --CF.sub.3, --OCH.sub.3, and --OCF.sub.3. In
some embodiments, substituted groups are substituted with one or
two of the preceding groups. In some embodiments, substituted
groups are substituted with one of the preceding groups. In some
embodiments, an optional substituent on an aliphatic carbon atom
(acyclic or cyclic, saturated or unsaturated carbon atoms,
excluding aromatic carbon atoms) includes oxo (.dbd.O).
The term "alkylene" refers to a divalent alkyl radical. Any of the
above mentioned monovalent alkyl groups may be an alkylene by
abstraction of a second hydrogen atom from the alkyl. In one
aspect, an alkelene is a C.sub.1-C.sub.6alkylene. In another
aspect, an alkylene is a C.sub.1-C.sub.4alkylene. Typical alkylene
groups include, but are not limited to, --CH.sub.2--,
--CH(CH.sub.3)--, --C(CH.sub.3).sub.2--, --CH.sub.2CH.sub.2--,
--CH.sub.2CH(CH.sub.3)--, --CH.sub.2C(CH.sub.3).sub.2--,
--CH.sub.2CH.sub.2CH.sub.2--, --CH.sub.2CH.sub.2CH.sub.2CH.sub.2--,
and the like.
The term "alkenyl" refers to an unsaturated aliphatic group having
at least one carbon-carbon double bond. In some embodiments,
C.sub.2-C.sub.8alkenyl has at least two carbons and up to 8 carbons
in said unsaturated aliphatic group.
The term "alkynyl" refers to an unsaturated aliphatic group having
at least one carbon-carbon triple bond. In some embodiments,
C.sub.2-C.sub.8alkynyl has at least two carbons and up to 8 carbons
in said unsaturated aliphatic group.
EXAMPLE
Example 1: Preparation of Exemplary Amine Derivative Anticancer
Agents
##STR00016##
Scheme 1. Preparation of Amine Derivative Anti-Cancer Agent from
4-hydroxy-2,3-dimethoxy-6-methyl-5-(3,7,11-trimethyldodeca-2,6,10-trienyl-
)cyclohex-2-enone
Exemplary anticancer agents were prepared according to Scheme 1
from
4-hydroxy-2,3-dimethoxy-6-methyl-5-(3,7,11-trimethyldodeca-2,6,10-trienyl-
)cyclohex-2-enone (100 mg, 0256 mmol) with a suitable amine
reagent. A more detailed experimental procedure for preparation of
Compound 1 is provided below.
##STR00017##
Compound 1:
(4S,5S,6S)-3-amino-4-hydroxy-2-methoxy-6-methyl-5-((2E,6E)-3,7,11-trimeth-
yldodeca-2,6,10-trien-1-yl)cyclohex-2-enone
A 30% of ammonium hydroxide (NH.sub.4OH) solution (2.6 mL, 66.8
mmol) was added to a solution of
4-hydroxy-2,3-dimethoxy-6-methyl-5-(3,7,11-trimethyldodeca-2,6,10-trienyl-
)cyclohex-2-enone (100 mg, 0.256 mmol) in MeOH (26 mL) under
N.sub.2 at ice bath. The mixture was stirred at room temperature
under N.sub.2 for 16 h. The excess ammonium hydroxide was removed,
and the residue was extracted with CH.sub.2Cl.sub.2 (10
mL.times.3). The combined organic phases was washed with brine 10
mL, dried over Na.sub.2SO.sub.4, and concentrated in vacuo to get
product 64 mg (0.17 mmol, 67%). .sup.1H (600 MHz; CDCl.sub.3)
.delta. 1.14 (3H, d, J=7.4 Hz), 1.54 (3H, s), 1.54-1.56 (br, 6H),
1.63 (3H, s), 1.89-1.98 (5H, m), 1.98-2.07 (5H, m), 2.31-2.38 (1H,
m), 2.39-2.46 (1H, m), 3.59 (3H, s), 4.72 (1H, d, J=4.4 Hz),
5.02-5.06 (2H, m), 5.10 (1H, t, J=7.3 Hz); .sup.13C (125 MHz;
CDCl.sub.3) .delta. 15.9, 16.0, 16.1, 17.6, 25.6, 26.0, 26.5, 26.7,
39.6, 39.8, 42.0, 46.0, 59.0, 66.0, 122.0, 123.9, 124.3, 128.9,
131.2, 135.1, 137.8, 154.8, 192.0; EI-MS, m/z 398 [M+Na].sup.+.
Compounds 2 to 11 were prepared similarly and confirmed with
analytical methods. The reagents and conditions are listed below in
Table 1 for the preparation of Compounds 1-11.
TABLE-US-00001 TABLE 1 Preparation of Compounds 1-11 with 3-amino
derivatives. Com- pound Temp. MS Yield No. RNH.sub.2 (.degree. C.)
Result (%) 1 NH.sub.4OH rt [M + Na].sup.+ 398 67 2 ##STR00018## rt
[M + H].sup.+ 404.19 [M + Na].sup.+ 426.13 >95 3 ##STR00019##
150 in sealed tube [[M + Na].sup.+ 480.25 12 4 ##STR00020## rt [M +
H].sup.+ 404.13 [M + Na].sup.+ 426.13 45 5 ##STR00021## rt [M +
H].sup.+ 433.28 [M + Na].sup.+ 455.17 >95 6 ##STR00022## rt [M +
H].sup.+ 461.25 [M + Na].sup.+ 483.32 99 7 ##STR00023## rt [M +
Na].sup.+ 442.12 47 8 ##STR00024## rt [M + Na].sup.+ 488.15 43 9
##STR00025## 150 in sealed tube [M + Na].sup.+ 489.09 52 10
##STR00026## 150 in sealed tube [M + Na].sup.+ 494.17 83 11
##STR00027## rt [M + Na].sup.+ 541.14 83
The following non exclusive exemplary compounds are prepared
accordingly.
##STR00028## ##STR00029## ##STR00030## ##STR00031##
##STR00032##
Example 2: MTT Assay
The MTT (3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium
bromide) cell viability assay is a colorimetric assay system, which
measures the reduction of a tetrazolium component (MTT) into an
insoluble blue/purple colored formazan product by succinate
tetrazolium reductase in mitochondria of viable cells. The
absorbance of the complex is read spectrophotometrically and is
directly proportional to the number of live or viable cells.
Formazan formation can therefore be used to assess and determine
the survival rate of cells.
Human hepatoma cell lines (HepG2) were obtained from American Type
Culture Collection (Rockville, Md., USA) and cultured in Minimum
Essential Medium Alpha (Invitrogen/Gibco BRL, Grand Island, N.Y.,
USA) at 37.degree. C. in 5% CO.sub.2 in culture media supplemented
with 10% fetal bovine serum (FBS) (Invitrogen/Gibco BRL) and 100
U/ml streptomycin and penicillin (Invitrogen/Gibco BRL).
100 .mu.L of HepG2 cells (1.times.10.sup.4 cells per well) were
seeded in 96 well plates and pre-incubated. After 24 h, the cells
were exposed to various concentrations of Compounds 1-11, and the
known compound
4-hydroxy-2,3-dimethoxy-6-methyl-5-(3,7,11-trimethyldodeca-2,6,10-trienyl-
)cyclohex-2-enone (Compound S) in a volume of 200 .mu.L for 48 h.
20 .mu.L of MTT reagent (5 mg/mL) was added to each well and then
incubated in 5% CO.sub.2 at 37.degree. C. for 4 h. The media was
replaced with 200 .mu.L of DMSO to dissolve the MTT tetrazolium
crystal. Absorbance was measured at 570 nm using a microplate
reader. The results are shown in Table 2.
TABLE-US-00002 TABLE 2 MTT Assay Results Compound No. IC.sub.50
(ug/ml) 1 0.13 2 -- 3 -- 4 0.18 5 0.87 6 -- 7 1.95 8 -- 9 -- 10
0.91 11 2.29 S 2.4
The data clearly shows the exemplary amine derivatized anticancer
agents exhibit unexpectedly better inhibition compared to
4-hydroxy-2,3-dimethoxy-6-methyl-5-(3,7,11-trimethyldodeca-2,6,10-trienyl-
)cyclohex-2-enone against liver cancer cells with some of them 10
to 20 folds better. In particular, Compounds 1, 4, 5, and 10
exhibit much better unexpected inhibition against liver cancer
cells. With the major difference between
Example 3: Efficacy Test of Exemplary Compound 1 on Liver Cancer
Xenograft Model
Thymic nude mice (National Laboratory Animal Center) of four to six
weeks old were used and maintained in laminar flow cabinets under
pathogen-free condition. Human HCC Mahlavu cells (1.times.10.sup.6
cells) carrying luciferase gene were re-suspended in 20 .mu.L of
PBS containing 50% matrigel (BD Biosciences, MA) and injected into
the left lateral liver lobe of athymic nude mice (National
Laboratory Animal Center) with 27-gauge needle by sterile
techniques (Lu et al., 2007). For bioluminescence imaging, the
animals were injected with the luciferase substrate D-luciferin at
a dose of 150 mg/kg in 0.2 mL sterile isotonic saline.
Tumor growth and metastatic status was monitored using the Xenogen
International Veterinary Information Service (IVIS) imaging system
every two weeks. All images were obtained after intraperitoneal
injection of luciferin (100 mg/kg body weight; Synchem, Elk Grove
Village, Ill., United States). Ten minutes after injection of
luciferin, nude mice were placed onto the Xenogen IVIS imaging
stage and were continuously sedated during image acquisition. Image
analysis and bioluminescence quantification were performed using
Living Image software (Caliper Life Sciences, Hopkinton, Mass.,
United States). BLI is based on the detection of light emitted by
living cells expressing a luciferase gene. The tumor-bearing mice
were treated with vehicle (olive oil) or Compound 1 at 120 mg/kg by
oral gavage twice per day and 5 days per week for 4 weeks. The body
weight was measured twice weekly.
Results
FIG. 1 shows decrease of the tumor size of THP-1 xenograft mice
treated with the exemplary test compound. The tumor-bearing mice
were treated with vehicle (olive oil) or Compound 1 at 120 mg/kg by
oral gavage twice per day and 5 days per week for 4 weeks. The
tumor volume was measured twice weekly. ***P<0.005 compared to
vehicle control.
FIG. 2 shows the decrease tumor mass weight at the end point of
THP-1 xenograft mice treated with test compound. At the end of week
12, tumor mass of the mice treated with Compound 1 decrease at
least 4 folds compared with one without treatment (the
control).
FIG. 3 shows the results of body weight changes of THP-1 xenograft
mice treated with or without test compound. The almost no change of
body weight indicates the test compound did not cause any serious
side effects.
These results clearly demonstrate the effectness and efficacy
exemplary invention Compound 1 on liver cancer treatment based on
xenograft model.
While preferred embodiments of the present invention have been
shown and described herein, it will be obvious to those skilled in
the art that such embodiments are provided by way of example only.
Numerous variations, changes, and substitutions will now occur to
those skilled in the art without departing from the invention. It
should be understood that various alternatives to the embodiments
of the invention described herein may be employed in practicing the
invention. It is intended that the following claims define the
scope of the invention and that methods and structures within the
scope of these claims and their equivalents be covered thereby.
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